1. Field of the Invention
The present invention relates to a protein, named ACA1, purified from Antrodia camphorata, and more particularly to the amino acid sequence of ACA1 and its immunomodulatory activity, as demonstrated by its stimulatory activity toward RAW 264.7 macrophages and mouse splenocytes.
2. Description of the Related Art
Antrodia camphoratais is a Taiwan specific mushroom having a special regional value. Although it has been existing in Taiwan for a long time, its first research report was published only as recently as in 1990. Antrodia camphoratais classified as Mycota, Basidiomycotina, Hymenomycetes, Phyllophorales, Polyporaceae, Antrodoa. The carpophores of Antrodia camphorata are perennial having a strong camphor tree aroma. Its shape is plate form or bell form, very different from the normal shape of ganoderma. The plate form, it cap is orange (yellow) color and full of gills, its floor plate has ivory color phellem, the Antrodia camphorata adhering to the inside wall of hollow wood of cinnamomum comphora grows by said phellem. The bell form, its carpophore (bell cap) also is orange (yellow) color and full of gills (4-5 gills/mm), and it has spores with bitter flavor. It is orange color in fresh, and then becomes dark orange color or brown color, bell body shows blackish green shell. To observe basidiospore by microscope, its shape is smooth, colorless, semi-bended pillar (3.5-5.0×1.5-2 μm).
There had been few reports on the physiological function of Antrodia camphoratabefore 2002, and there also lack the clinical and pharmaceutical evidences from science research. The well-known physiological functions of Antrodia camphorata are the curative effects passed on folk therapy, therefore in-depth research using scientic experiment is needed. On the physiological of fungus, Antrodia camphorata is the only mushroom that can metabolize a large number of bacteriostasis safrole of cinnamomum comphora and grows normally. It shows that Antrodia camphorata has a physiological metabolism function different from other mushrooms. The research, Cheng Yi Hua, 1994, indicates all of the triterpenoids elements of carpophore of Antrodia camphorata having 24(28)-en frame on its branch. It shows that the titerpenoids metabolism of Antrodia camphorata is very different from ganoderma, these branch frames can be the characteristic elements of Antrodia camphorata, the 30% methanol extract of Antrodia camphoratais more than 3% methanol extract of ganoderma, and Antrodia camphorata is bitterer than Ganoderma, because of Antrodia camphorata has more oxidated triterpenoids and sterol. Wang Bo-Che, food industry, 1998, 30: 1-36., indicates that Antrodia camphorata has valuable evaluation for detoxification of food poisoning diarrheabellyache vomit pesticide. Antrodia camphorata is very well done in treating of hepato tumor and uterus tumor, settling spirit, and it is also a magic tool for protecting liver and dissolving drunkenness.
Recently, the academia has begun to actively research the functions and active ingredients of Antrodia camphorata. Tsai Yan Hui, 2002, master thesis of Department of Food Science, National Chung Hsing University, indicates that triterpene compound β which isolated from Antrodia camphorata carpophore has reducing acute liver damage induced by carbon tetrachloride in the rat liver functional experiments and reducing mouse's blood GPT value. Song, T. Y et al., 2002, Journal Agriculture Food Chemistry, 50: 3322-3327 and 2003, Journal Agriculture Food Chemistry, 51: 1571-1577, indicate that the ferment filtrate of Antrodia camphorata has highly ability to protect acute liver damage of rat induced by carbon tetrachloride. Chen Li Yan et al., 2002, in seminar and Chen Xin Yi, 2002, master thesis of China Medical School Institute of Nutrition, indicate adequate ferment filtrate of Antrodia camphorata mycelium has positive effect for liver physiological function, it can reduces liver fibrosis and denature, increases GSH content in liver and erythrocyte and the activity of antioxidate enzyme, decreases liver lipid peroxidation, and good for decreases body oxidation pressure. Yang Su We, 1991, master thesis of National Taiwan University Institute of Pharmacy, indicates after extract of Antrodia camphorata divide extracted with ethyl acetate and water, the activities of anticholine and antiserotonin of ethyl acetate level are the strongest, and have the curative effect certainly. Chen Jin Chu, 2001, in seminar, discloses the preliminary safe experiment of Antrodia camphorata, he detects that Antrodia camphoratamycelium has no toxicity for various cell and mouse, and it is need to systematicness and widespread study the immunity regulationsanti-tumors and other physiologival functions of Antrodia camphorata. Chen Quing Nong et al., 1999, in seminar, indicate the special aroma of Antrodia camphorata has the ability of antibiotic and antioxidation, and it can apply in aromatherapy, the results of appraise said aroma elements are perpene alcohols and ethyl hexadecanoate etc. Grape King Inc. performs study of the ability of antihepatitis B virus and immunity of Antrodia camphorata, it shows rude extract of Antrodia camphorata has stronger activity of antihepatitis B virus, and after concanavalin A (ConA) treats, the Antrodia camphorata mycelium can promotes lymphocyte proliferation and irritates spleen cell producing cytokine IL-2.
In mushroom increasing immunity way, Lee S. S. et al., 1995, Journal of Chinese Medicine, 6:1-12, indicate every extracts of mushroom can increase immune cell activity, and promote cytokine and interferon production. Chen W. C. et al., 1999, the American journal of Chinese medicine, indicate the extract of mushroom can restore the immunity of mouse which immunity decreasing after irradiation. Furthermore, Ding Huai Qian, 2000, Food industry, 32:28-42, indicates scholars had been study in activity of immune physiology of edible and medicinal using mushrooms, such as ganodermacoridus ganodermatremellachinese caterpillar fungushericium erinaceumpolyporusgolden mushroommushroom and maitake mushroom, base on different kinds of mushroom, individual of them possesses some abilities of promoting lymphocyte proliferation, activating B cell and increasing serum antibody content, raising phagocytic ability of macrophage and nature killer cell, promoting reaction of SRBC and PFC, having inhibitions to resist immunity inhibitor such as cyclophosphamide, mercaptopurine, fluorouracil, and promoting ConA induced mouse spleen lymphocyte proliferation, isoform antigen irritating transformation of lymphocyte. Lieu C.W. et al., 1992, Anticancer Research, 12:1211-1215Sakagami MRS. et al., 1993, Anticancer Research, 13:671-675Wang H. et al., 2001, Biochemical and biophysical research communications, 275: 810-816 and 2002, Biochemical and biophysical research communications, 289: 750-755, organism activates immune system to kill cancer cell or inhibit tumor growing due to the extract of mushroom can activate various body immune cell raises the phagocytic ability of macrophage and nature killer cellstimulates immune cell secreting various interferon and cytokines to inhibit tumor cell. Therefore, it is positive relation in the ability of anti-tumor of the extract of mushroom and the ability of promoting immunity of the extract of mushroom.
Antrodia camphorata is a Taiwan specific fungus, and has various physiology activities, but its activated elements are not clear before. The protein ACA1 from Antrodia camphorata mycelium was purified and its immunomodulatory activity disclosed in this invention.